Adult High-grade Gliomas Research Articles

HGG-29. A CASE OF CIRCUMSCRIBED HIGH-GRADE ASTROCYTOMA WITH ATRX AND CDKN2A/B ALTERNATIONS WHO WAS INITIALLY DIAGNOSED AS GLIOBLASTOMA AND HAS 20 YEARS SURVIVAL

Pediatric high-grade gliomas are rare and often hard to classify, which grow locally and show longer survival than diffuse high-grade gliomas in adults. We report a case of circumscribed high-grade astrocytoma who was initially diagnosed as glioblastoma and has 20 years survival. A 7-year-old girl suffered from epileptic seizure due to a left occipital lobe tumor. The tumor was resected in another hospital and diagnosed as glioblastoma. The tumor disappeared after extended local irradiation and chemotherapy using nimustine hydrochloride (ACNU) and cisplatin (CDDP). Eighteen years after initial onset, first recurrence was confirmed as the intra-tumoral hemorrhage. The tumor was resected and diagnosed as anaplastic oligoastrocytoma. After 6 courses of temozolomide (TMZ), the tumor disappeared. Twenty years after initial onset, the second local recurrence was confirmed. Although gamma knife and TMZ was performed, the tumor did not disappear. The tumor was surgically resected. Histopathology showed localized growth with some infiltration and mitosis but lacked pseudopallisading and microvascular proliferation. The tumor was diagnosed as circumscribed high-grade astrocytoma. Immunostaining revealed ATRX nuclear loss and CDKN2A / B homozygous deletion. After 10 courses of TMZ, the third local recurrence was confirmed. The tumor was completely removed and has not occurred recurrence more than 3 months after the last operation. Circumscribed high-grade glioma is expected to survive longer than invasive glioma. Pediatric gliomas should differ from adult gliomas in the genes of tumorigenesis. Care should be taken for its diagnosis and treatments. We also need a new classification based on histology and gene profile. HGG-30, ANALYSIS OF PEDIATRIC GLIOMAS IN OUR INSTITUTE Kaoru Tamura, Mai Fujioka, Masae Kuroha, Motoki Inaji, Yoji Tanaka, Tadashi Nariai, and Taketoshi Maehara; Tokyo Medical and Dental University, Tokyo, Japan. PURPOSE: Recent advances in genetic interrogation of pediatric glioma increase the importance of molecular diagnosis using surgical specimen. However, surgical resection may be avoided to preserve quality of life, especially in brain stem glioma cases. We retrospectively examined diagnosis and treatment of pediatric gliomas in our hospital. METHODS: This study includes 14 consecutive glioma patients under the age of 18 who underwent initial treatment at our hospital from 2000 to 2019. Histopathological diagnosis, clinical course and molecular status such as IDH, H3F3A and BRAF were analyzed. RESULTS: 5 patients (1 pilocytic astrocytoma (PA), 3 diffuse astrocytomas, 1 oligodendroglioma were treated only by surgical resection (group A). 7 patients (1 PA, 1 anaplastic oligodendroglioma, 2 diffuse midline gliomas and 3 glioblastomas (GBM)) received radiation and/or chemotherapy after surgical resection (group B). 2 diffuse intrinsic pontine gliomas (DIPG) received radiation and chemotherapy without surgical resection (Group C). No IDH mutation was observed in all pathological specimen obtained cases. BRAF alteration was observed in all PA cases. 1 case of GBM had BRAF V600Emutation and the other had H3K27M mutation. During a median of 7.7 years of follow-up, group A patients have no recurrence. Group B includes various diagnosis and prognosis. 2 group C patients diagnosed DIPG by MRI showed different clinical courses. CONCLUSION: Pediatric gliomas include diverse biological subgroups and show broad range of clinical behavior. Since pediatric glioma has a low incidence and a wide variety of genetic mutations, multicenter study is important to improve the treatment of pediatric glioma.

DIPG-68. ALPHA-THALASSEMIA X-LINKED MENTAL RETARDATION PROTEIN (ATRX) LOSS-OF-FUNCTION IN A MOUSE MODEL OF DIFFUSE INTRINSIC PONTINE GLIOMA

Diffuse Intrinsic Potine Glioma (DIPG) is a rare pediatric brain tumor for which no cure or efficacious therapies exist. Previous discoveries have revealed that, DIPG harbors distinct genetic alterations, when compared with adult high-grade glioma (HGG) or even with non-DIPG pediatric HGGs. ATRX alteration is found in 9% of clinical cases of DIPG, and significantly overlaps with H3.3K27M mutation and p53 loss, the two most common genetic changes in DIPG, found in 80% and 77% clinical cases, respectively. Here we developed genetically engineered mouse model of brainstem glioma using the RCAS-Tv-a system by targeting PDGF-B overexpression, p53 loss, H3.3K27M mutation and ATRX loss-of function to Nestin-expression brainstem progenitor cells of the neonatal mouse. Specifically, we used Nestin-Tv-a; p53 floxed; ATRX heterozygous female and Nestin-Tv-a; p53 floxed; ATRX floxed male breeders, generated offsprings with ATRX loss of function (n=18), ATRX heterozygous females (n=6), and ATRX WT (n=10). Median survial of the three groups are 65 days, 88 days and 51 days, respectively. Also, ATRX null mice is lower in tumor incidence (44.4%), compared with ATRX WT (80%). We evaluated the pathological features of DIPG with or without ATRX alteration, RNA-seq is performed to identify differentially expressed genes between ATRX WT and loss-of-function. In conclution, this study generated the first genetically modified mouse model studying ATRX loss-of-function in DIPG, and suggested that ATRX loss-of-function in DIPG may slow down tumorigenesis and decrease tumor incidence.

HGG-11. HIGH-GRADE GLIOMAS IN ADOLESCENTS AND YOUNG ADULTS HIGHLIGHT HISTOMOLECULAR DIFFERENCES WITH THEIR ADULT AND PAEDIATRIC COUNTERPARTS

BACKGROUNDConsidering that paediatric high-grade gliomas (HGGs) are biologically distinct from their adult counterparts, the objective of this study was to define the landscape of HGGs in adolescents and young adults (AYAs).METHODSWe performed a multicentric retrospective study of 112 AYAs from adult and paediatric Ile-de-France neurosurgical units, treated between 1998 and 2013 to analyse their clinicoradiological and histomolecular profiles. The inclusion criteria were age between 15 and 25-years, histopathological HGG diagnosis, available clinical data, pre-operative and follow-up MRI. MRI and tumoral samples were centrally reviewed. Immunohistochemistry and complementary molecular techniques such as targeted/next generation sequencing, whole exome sequencing and DNA-methylation analyses were performed to achieve an integrated diagnosis according to the 2016 WHO classification.RESULTSBased on 80 documented AYA patients, HGGs constitute heterogeneous clinicopathological and molecular groups, with a predominant representation of paediatric-subtypes (Histone H3-mutants, 40%) but also adult-subtypes (IDH-mutants, 28%) characterized by the rarity of oligodendrogliomas, IDH-mutant and 1p/19q co-deleted and the relative high frequency of “rare adult IDH mutations” (20%). H3G34-mutants (14%) represent the most specific subgroup in AYAs. In the H3K27-mutant subgroup, the non-brainstem diffuse midline gliomas are more frequent (66.7%) than diffuse intrinsic pontine gliomas (23.8%), contrary to children. We found that WHO grade has no prognostic value, but molecular subgrouping has major prognostic importance.CONCLUSIONSHGGs in AYAs could benefit from a more personalized neuro-oncological management, driven by molecular subtyping rather than age group. Collaborative efforts are needed from paediatric and adult neuro-oncology teams to improve the management of HGGs in AYAs.

The Use of 5-Aminolevulinic Acid to Assist Gross Total Resection of Paediatric Posterior Fossa Tumours

Background: 5-aminolevulinic acid (5-ALA) use is well established in the resection of adult high-grade gliomas. There is growing interest in its usefulness in the paediatric population. The potential benefit of 5-ALA-guided resection motivated our unit to offer the established adult protocol as off-label use. Objective: to determine if 5-ALA guided resection was routinely useful and offered increased gross total resection (GTR) results. Methods: Nineteen patients harbouring a posterior fossa tumour suggestive of either an ependymoma or medulloblastoma (MB) underwent surgery between January 2018 and October 2019. The mean age was 5 years (range 2–12 years). A dose of 20 mg/kg of 5-ALA (Gliolan®) was given 4 h preoperatively. Intraoperatively, the tumours were viewed under violet-blue light and the presence of fluorescence was recorded. Fluorescence status was compared with histopathological classification and grade, Ki-67 index, GTR rate, and a subjective determination of “usefulness” was determined. Results: The case series included ependymoma grade II (n = 6), ependymoma grade III (n = 4), and MB grade IV (n = 9). For the combined cohort, the strong fluorescence rate was 68% (n = 13), the heterogenous fluorescence rate was 26% (n = 5), and the completely negative fluorescence rate was 5% (n = 1). The strong fluorescence rate of 90% found in the combined ependymoma group compared to the 45% strong fluorescence rate in the MB group was statistically significant (p = 0.05). Within the MB group the Ki-67 index was found to be significantly higher in the strongly fluorescent group as opposed to the patchy or non-fluorescent group (77.5 vs. 40%, p = 0.016). Fluorescence was determined to be useful in 63% of all cases. There was no significant relationship between fluorescence and GTR. The relationship between perceived usefulness and resection was not statistically significant. No adverse drug reactions were recorded. Conclusion: This case series adds to the growing body of evidence demonstrating the safety of 5-ALA in the paediatric population. 5-ALA guided resection was found to be useful in the majority of cases but this did not correlate with GTR status. Ependymomas reliably fluoresce in 90% of cases, and 5-ALA-guided resection should be considered when a preoperative diagnosis of ependymoma is suspected.

Adult diffuse midline gliomas: Clinical, radiological, and genetic characteristics

Diffuse midline gliomas (DMGs) are a diffuse glioma subtype arising from midline brain structures. It is predominantly a disease of childhood; however, it can also occur in adults. Adult DMG has not been previously well described. The aim of this study was to define the characteristics of adult DMG. We described and analyzed the clinical, radiological, and genetic alterations of 9 adult DMGs and compared them with those of 257 non-midline adult high-grade IDH-WT gliomas. The median age of all patients was 38-years old (23–68-years). Most common symptoms were headache, motor/sensory deficit, ataxia, cranial nerve deficit, and confusion. Tumor locations were brainstem (44.5%), thalamus (22.2%), pineal region (22.2%), spinal cord (22.2%), and cerebellum (11.1%). Six-patients (66.7%) were H3 K27M-WT and three (33.3%) were H3 K27M-mutant. In addition to H3 K27M mutations, TP53 gene (55.5%), CDKN2A/B and TERTp (33.3%), PDGFRA (33.3%), PIK3CA, PTEN, KDR, NF1, and MYC (22.2%) were the most frequently mutated genes. Neither IDH1/IDH2 nor EGFR alterations were present. Compared to non-midline high-grade glioma, adult DMG patients were younger (38 vs 61 years, p < 0.001) and lacked EGFR-alterations (0/9 vs 123/257, p = 0.004). The median survival of DMG and non-midline high-grade gliomas was 19 and 18 months respectively (p = 0.964). Our data support that adult DMGs have different oncogenic drivers compared to non-midline high-grade gliomas. Regardless of H3 K27M mutation status, neither of the nine adult DMG cases demonstrated IDH1/IDH2 or EGFR alterations. Larger multi-institutional studies are needed to further characterize the biology of this rare type of diffuse glioma in adults.

Everolimus improves the efficacy of dasatinib in PDGFRα-driven glioma.

Pediatric and adult high-grade gliomas (HGGs) frequently harbor PDGFRA alterations. We hypothesized that cotreatment with everolimus may improve the efficacy of dasatinib in PDGFRα-driven glioma through combinatorial synergism and increased tumor accumulation of dasatinib. We performed dose-response, synergism, P-glycoprotein inhibition, and pharmacokinetic studies in in vitro and in vivo human and mouse models of HGG. Six patients with recurrent PDGFRα-driven glioma were treated with dasatinib and everolimus. We found that dasatinib effectively inhibited the proliferation of mouse and human primary HGG cells with a variety of PDGFRA alterations. Dasatinib exhibited synergy with everolimus in the treatment of HGG cells at low nanomolar concentrations of both agents, with a reduction in mTOR signaling that persisted after dasatinib treatment alone. Prolonged exposure to everolimus significantly improved the CNS retention of dasatinib and extended the survival of PPK tumor-bearing mice (mutant TP53, mutant PDGFRA, H3K27M). Six pediatric patients with glioma tolerated this combination without significant adverse events, and 4 patients with recurrent disease (n = 4) had a median overall survival of 8.5 months. Our results show that the efficacy of dasatinib treatment of PDGFRα-driven HGG was enhanced with everolimus and suggest a promising route for improving targeted therapy for this patient population.

Abstract 4943: The stem cell factor LIN28B regulates proliferation and apoptosis in diffuse intrinsic pontine glioma

Abstract Diffuse Intrinsic Pontine Glioma (DIPG) is a lethal pediatric brain tumor. With radiation therapy as the primary treatment, the median survival after diagnosis is 6 months. Identifying molecular markers that regulate tumor growth is urgently needed to develop targeted therapies. LIN28B is a stem cell factor expressed during normal neural development and re-expressed in cancer cells. In many cancers, LIN28B acts as an RNA-binding protein and downregulates the tumor-suppressing let-7 microRNAs. We had previously shown that LIN28A, a homolog of LIN28B, regulates invasion and tumorigenicity in adult high-grade gliomas. LIN28B is known to be highly expressed in DIPG. Thus, we hypothesized that LIN28B would be important in DIPG by promoting tumor growth. We observed that LIN28B was expressed in patient-derived DIPG cell lines (JHHDIPG1, JHHDIPG16A, SUDIPGXIII, HSJD-007, SF7761). Knockdown of LIN28B using short hairpin RNAs decreased DIPG cell proliferation (BrdU incorporation) between 50% to 90% (p&amp;lt;0.01). The knockdown of LIN28B also increased DIPG cell apoptosis (cleaved caspase-3 and cleaved PARP expression) by 2- to 3-fold (p&amp;lt;0.01). To determine the molecular mechanism of LIN28B-mediated phenotypes in DIPG, we studied canonical downstream effectors of LIN28B. We found that when LIN28B was suppressed in DIPG cells, there was a decreased expression of the DNA-binding oncoprotein called high mobility group AT-hook 2. These findings suggest that LIN28B could be a potential therapeutic target for this devastating pediatric brain tumor. Future studies will focus on validating the role of LIN28B in DIPG growth in vivo using orthotopic zebrafish and mouse models. Citation Format: Huizi Guo, Harpreet Kaur, Charles G. Eberhart, Eric H. Raabe. The stem cell factor LIN28B regulates proliferation and apoptosis in diffuse intrinsic pontine glioma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4943.

Prognostic Nomograms for Primary High-Grade Glioma Patients in Adult: A Retrospective Study Based on the SEER Database.

Purpose In our study, we aimed to screen the risk factors that affect overall survival (OS) and cancer-specific survival (CSS) in adult glioma patients and to develop and evaluate nomograms. Methods Primary high-grade gliomas patients being retrieved from the surveillance, epidemiology and end results (SEER) database, between 2004 and 2015, then they randomly assigned to a training group and a validation group. Univariate and multivariate Cox analysis models were used to choose the variables significantly correlated with the prognosis of high-grade glioma patients. And these variables were used to construct the nomograms. Next, concordance index (C-index), calibration plot and receiver operating characteristics (ROCs) curve were used to evaluate the accuracy of the nomogram model. In addition, the decision curve analysis (DCA) was used to analyze the benefit of nomogram and prognostic indicators commonly used in clinical practice. Results A total of 6395 confirmed glioma patients were selected from the SEER database, divided into training set (n =3166) and validation set (n =3229). Age at diagnosis, tumor grade, tumor size, histological type, surgical type, radiotherapy and chemotherapy were screened out by Cox analysis model. For OS nomogram, the C-index of the training set was 0.741 (95% CI: 0.751-0.731), and the validation set was 0.738 (95% CI: 0.748-0.728). For CSS nomogram, the C-index of the training set was 0.739 (95% CI: 0.749-0.729), and the validation set was 0.738 (95% CI: 0.748-0.728). The net benefit and net reduction in inverventions of nomograms in the decision curve analysis (DCA) was higher than histological type. Conclusions We developed nomograms to predict 3- and 5-year OS rates and 3- and 5-year CSS rates in adult high-grade glioma patients. Both the training set and the validation set showed good calibration and validation, indicating the clinical applicability of the nomogram and good predictive results.

Abstract IA20: Neuronal activity promotes proliferation of normal and neoplastic glial cells

Abstract Neuronal activity regulates the proliferation and differentiation of oligodendrocyte precursor cells during development and in adulthood. In the healthy brain, this results in activity-regulated plasticity of myelination and subsequent modulation of neural circuit function evident in oligodendrogenesis-dependent behavioral changes. The robust mitogenic effect of neuronal activity on normal neural precursor and oligodendroglial precursor cells, a putative cellular origin of pediatric high-grade gliomas (HGG), suggests that dysregulated or “hijacked” mechanisms of myelin plasticity might similarly promote proliferation in this devastating group of childhood brain cancers. Using in vivo and in situ optogenetic techniques together with patient-derived high-grade glioma cell cultures and xenograft models, we have demonstrated that neuronal activity similarly promotes proliferation and growth of both pediatric and adult high-grade glioma subtypes. Crucial mechanisms mediating activity-regulated high-grade glioma growth include secretion of Brain Derived Neurotrophic Factor (BDNF) and the synaptic protein neuroligin-3 (NLGN3), which induces multiple oncogenic signaling pathways together with robust changes in synaptic gene expression in glioma cells. Nlgn3 is necessary for the growth of high-grade glioma xenografts in the mouse brain, and NLGN3 expression levels in human HGG negatively correlate with patient overall survival. Thus, neuronal activity not only modulates the structure and function of the brain’s myelinated infrastructure, but neurons also play an important role in the brain tumor microenvironment, with activity-regulated secretion of NLGN3 emerging as an unexpected and therapeutically targetable mechanism underlying axon-glioma interactions and promoting neuronal activity-regulated cancer growth.

A systematic review of tumor treating fields therapy for high-grade gliomas.

Tumor treating fields (TTF) is a unique treatment modality that utilizes alternating electric fields to deliver therapy. Treatment effects have been assessed in patients with newly diagnosed and recurrent glioblastoma in clinical trials and retrospective studies. While the results of these studies led to FDA approval for both populations, a portion of the neuro-oncology and neurosurgery community remains skeptical of TTF. Thus, this review aims to systematically summarize and evaluate prior studies investigating the efficacy and safety of TTF in patients with high-grade gliomas. A systematic review of the literature was performed according to PRISMA guidelines from database inception through February 2019. To be included, studies must have investigated the efficacy of TTF in adult high-grade glioma patients. In total, 852 studies were initially identified, 9 of which met final inclusion criteria. In total, 1191 patients were identified who received TTF. Included studies consisted of two pilot clinical trials, two randomized clinical trials, and five retrospective studies. In randomized clinical trials, TTF improved survival for newly diagnosed glioblastoma patients but not for recurrent glioblastoma patients. Adverse skin reactions were the primary adverse effect associated with TTF. While TTF has been evaluated for safety and efficacy in a number of studies, concerns remain regarding study design, quality of life, and cost of therapy. Further investigation is needed regarding the therapy, and ongoing trials are already underway to provide more data regarding therapy outcomes and interactions in combination regimens.

A predictive analysis approach for paediatric and adult high-grade glioma: miRNAs and network insight.

BackgroundBrain tumours are the most common solid tumour in children and are a cause of mortality in adults. Most cases of brain tumour-related death are attributed to glioblastoma (GBM), with an elevated rate for high-grade glioma (HGG). Showing strong heterogeneity, the lesion location, molecule expression and type of HGG differ between adults and children. However, with regard to pathogenesis, brain tumours are expected to have the same underlying molecular processes.MethodsIn this study, we obtained data from the Gene Expression Omnibus (GEO) database to analyse molecular expression in HGG between adults and children. The same and different mutations were identified in these groups, and the genes involved were compared using Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. Molecular analysis revealed the same trend of differences between children and adults, which was verified in The Cancer Genome Atlas (TCGA).ResultsA total of 12 microarrays including 455 HGG patients were screened. Through a rigorous intersecting process, we identified miR-10a, miR-10b, and miR-139 as having common differences, as well as 6 target genes, such as CDK6, SOX4 and VEGFA, etc. And 12 long noncoding RNAs (lncRNAs).ConclusionsWe identified that these key molecules are involved in development and progression of HGG between adults and children. The findings provide a comprehensive description of the similarities in advanced diseases between adults and children and molecular diagnostic directions for precision small-molecule medicine to treat HGG in different age populations.

RAF and MEK inhibitor therapy in adult patients with brain tumors: a case-based overview and practical management of adverse events.

Targeted therapy has gained mainstream attention with notable successes against specific genetic mutations in many cancers. One particular mutation, the BRAF V600E mutation, is present in a small subset of gliomas in adults. Although clinical experience and trial data of RAF-targeted therapy in adults with glioma are lacking at this time, the poor prognosis of adult high-grade glioma has led neuro-oncology practitioners to consider the use of targeted therapy in these patients. In this manuscript, we describe the use of RAF and MEK inhibitors in adults with recurrent glioma. We discuss the utility of these agents, describe their toxicities, and give examples of management strategies. Given the significant toxicities of RAF and MEK inhibitors, along with the long potential duration of treatment, neuro-oncology providers should counsel patients carefully before initiating therapy and monitor them closely while undergoing treatment with RAF-targeted therapy.

High-grade gliomas in adolescents and young adults highlight histomolecular differences from their adult and pediatric counterparts

Considering that pediatric high-grade gliomas (HGGs) are biologically distinct from their adult counterparts, the objective of this study was to define the landscape of HGGs in adolescents and young adults (AYAs). We performed a multicentric retrospective study of 112 AYAs from adult and pediatric Ile-de-France neurosurgical units, treated between 1998 and 2013 to analyze their clinicoradiological and histomolecular profiles. The inclusion criteria were age between 15 and 25 years, histopathological HGG diagnosis, available clinical data, and preoperative and follow-up MRI. MRI and tumoral samples were centrally reviewed. Immunohistochemistry and complementary molecular techniques such as targeted/next-generation sequencing, whole exome sequencing, and DNA-methylation analyses were performed to achieve an integrated diagnosis according to the 2016 World Health Organization (WHO) classification. Based on 80 documented AYA patients, HGGs constitute heterogeneous clinicopathological and molecular groups, with a predominant representation of pediatric subtypes (histone H3-mutants, 40%) but also adult subtypes (isocitrate dehydrogenase [IDH] mutants, 28%) characterized by the rarity of oligodendrogliomas, IDH mutants, and 1p/19q codeletion and the relative high frequency of "rare adult IDH mutations" (20%). H3G34-mutants (14%) represent the most specific subgroup in AYAs. In the H3K27-mutant subgroup, non-brainstem diffuse midline gliomas are more frequent (66.7%) than diffuse intrinsic pontine gliomas (23.8%), contrary to what is observed in children. We found that WHO grade has no prognostic value, but molecular subgrouping has major prognostic importance. HGGs in AYAs could benefit from a specific classification, driven by molecular subtyping rather than age group. Collaborative efforts are needed from pediatric and adult neuro-oncology teams to improve the management of HGGs in AYAs.

Particle radiation therapy in the management of adult high-grade glioma: A narrative review

This narrative review summarizes the current status of the use of particle radiation therapy on the treatment of adult malignant gliomas. Due to the unique physical property, particle (e.g., proton or carbon-ion) beam radiation therapy can improve radiation dose distribution, thereby therapeutic radio for patients with brain malignancy. Therefore, particle beam radiation therapy is associated with low adverse events which have implications for improving quality of life for long-term survivors. In addition, there is a potential for safe dose escalation in selected patients. Malignant glioma is considered radioresistant; thus, particle beams of higher relative biological effectiveness, carbon-ion beam, for example, may further improve disease control in theory. Data on carbon-ion beam RT alone for the management of brain tumor are scarce. Most literature described proton beam alone or the use of combined proton/photon and carbon-ion beam boost for the treatment of glioma. Existing clinical evidence describes virtually no acute high-grade toxicities and limited late effects. Prospective clinical trials are needed to confirm the improved efficacy and favorable toxicity profile of particle beam radiation therapy on adult malignant glioma described in retrospective studies. Currently, multiple prospective trials are ongoing to answer such questions.

Proteasome inhibition as a therapeutic approach in atypical teratoid/rhabdoid tumors.

BackgroundAtypical teratoid/thabdoid tumor (AT/RT) remains a difficult-to-treat tumor with a 5-year overall survival rate of 15%–45%. Proteasome inhibition has recently been opened as an avenue for cancer treatment with the FDA approval of bortezomib (BTZ) in 2003 and carfilzomib (CFZ) in 2012. The aim of this study was to identify and characterize a pre-approved targeted therapy with potential for clinical trials in AT/RT.MethodsWe performed a drug screen using a panel of 134 FDA-approved drugs in 3 AT/RT cell lines. Follow-on in vitro studies used 6 cell lines and patient-derived short-term cultures to characterize selected drug interactions with AT/RT. In vivo efficacy was evaluated using patient derived xenografts in an intracranial murine model.ResultsBTZ and CFZ are highly effective in vitro, producing some of the strongest growth-inhibition responses of the evaluated 134-drug panel. Marizomib (MRZ), a proteasome inhibitor known to pass the blood–brain barrier (BBB), also strongly inhibits AT/RT proteasomes and generates rapid cell death at clinically achievable doses in established cell lines and freshly patient-derived tumor lines. MRZ also significantly extends survival in an intracranial mouse model of AT/RT.ConclusionsMRZ is a newer proteasome inhibitor that has been shown to cross the BBB and is already in phase II clinical trials for adult high-grade glioma (NCT NCT02330562 and NCT02903069). MRZ strongly inhibits AT/RT cell growth both in vitro and in vivo via a moderately well-characterized mechanism and has direct translational potential for patients with AT/RT.

Type 1 IGF receptor associates with adverse outcome and cellular radioresistance in paediatric high-grade glioma

High-grade glioma (HGG) is highly resistant to therapy, prompting us to investigate the contribution of insulin-like growth factor receptor (IGF-1R), linked with radioresistance in other cancers. IGF-1R immunohistochemistry in 305 adult HGG (aHGG) and 103 paediatric/young adult HGG (pHGG) cases revealed significant association with adverse survival in pHGG, with median survival of 13.5 vs 29 months for pHGGs with moderate/strong vs negative/weak IGF-1R (p = 0.011). Secondly, we tested IGF-1R inhibitor BMS-754807 in HGG cells, finding minimal radiosensitisation of 2/3 aHGG cell lines (dose enhancement ratios DERs < 1.60 at 2–8 Gy), and greater radiosensitisation of 2/2 pHGG cell lines (DERs ≤ 4.16). BMS-754807 did not influence radiation-induced apoptosis but perturbed the DNA damage response with altered induction/resolution of γH2AX, 53BP1 and RAD51 foci. These data indicate that IGF-1R promotes radioresistance in pHGG, potentially contributing to the association of IGF-1R with adverse outcome and suggesting IGF-1R as a candidate treatment target in pHGG.

BPTF regulates growth of adult and pediatric high-grade glioma through the MYC pathway.

High-grade gliomas (HGG) afflict both children and adults and respond poorly to current therapies. Epigenetic regulators have a role in gliomagenesis, but a broad, functional investigation of the impact and role of specific epigenetic targets has not been undertaken. Using a two-step, in vitro/in vivo epigenomic shRNA inhibition screen, we determine the chromatin remodeler BPTF to be a key regulator of adult HGG growth. We then demonstrate that BPTF knockdown decreases HGG growth in multiple pediatric HGG models as well. BPTF appears to regulate tumor growth through cell self-renewal maintenance, and BPTF knockdown leads these glial tumors toward more neuronal characteristics. BPTF’s impact on growth is mediated through positive effects on expression of MYC and MYC pathway targets. HDAC inhibitors synergize with BPTF knockdown against HGG growth. BPTF inhibition is a promising strategy to combat HGG through epigenetic regulation of the MYC oncogenic pathway.

PATH-53. IMMUNOLOGICAL PROFILING OF MUTATIONAL AND TRANSCRIPTIONAL SUBGROUPS IN PEDIATRIC AND ADULT HIGH-GRADE GLIOMAS

Abstract Immunological treatment strategies, including checkpoint inhibition, are currently under investigation for high-grade gliomas, but their success is limited. Hence, it is crucial to determine immunological pathways that can be targeted and to identify subgroups of patients likely to benefit from immunotherapies. Previous studies are still limited by comparably small sample sizes and there is only few data about specific immunological mechanisms in pediatric high-grade glioma. We gathered published gene expression data from 1135 adult and pediatric high-grade gliomas and applied a machine learning technique to determine their mutational (K27, G34, IDHmut, IDHwt) and transcriptional subtype. Subsequently, immune cell infiltration and functional immune pathway signatures were evaluated in correlation to histological diagnosis, age, transcriptional and mutational subtype. T-SNE analysis and unsupervised hierarchical clustering was applied to detect subgroup-specific immune microenvironments across all high-grade glioma subtypes. Four distinct microenvironmental phenotypes of immune cell infiltration were identified, which can be stratified into vascular, monocytic/stromal, monocytic/T-cell and APC/NK/T-cell dominant immune clusters. Immune cell infiltration correlated strongly with transcriptional and mutational subtypes but was independent of age and histological diagnosis (p&lt; 0.01). H3F3A mutated tumors had significantly fewer tumor infiltrating lymphocytes and macrophages. By including functional pathways and correlating the expression of immunostimulatory and inhibitory receptor/ligand interactions, we were able to define the immunological microenvironment and to identify possible immunological subtypes associated with poor prognosis as well as subtypes that might be especially amenable to checkpoint inhibition. In addition, comparisons of overall survival with the immunological microenvironment and specifically with immune checkpoint molecules revealed diverse correlations within the transcriptional and mutational subgroups. In conclusion, we shat that mutational and transcriptional subgroups of pediatric and adult high-grade gliomas are characterized by distinct immunological tumor microenvironments. Our analysis demonstrates the immunological heterogeneity within this entity and emphasizes an immune specific stratification of subgroups for upcoming immunotherapy trials.

PDTM-11. GAINING INSIGHTS INTO THE INFLAMMATORY MICROENVIRONMENT OF PEDIATRIC HIGH-GRADE GLIOMAS USING GEMMs AND PATIENT SAMPLES

Abstract Pediatric high-grade gliomas (pHGG) account for the most cancer-related deaths in children, as there are no effective therapies available. It is known tumor associated macrophages (TAM) can make up 30–40% of the total tumor cell mass in adult high-grade gliomas, promoting tumor growth and immune evasion. This raises the question of whether pHGGs possess a distinct constituency of TAMs due to their unique genetic and epigenetic landscapes. To uncover the composition and behavior of TAMs in pHGG we utilize RCAS/tva, a somatic cell-type specific gene transfer system which allows us to recapitulate all major subtypes of pHGG in newborn immunocompetent mice, including histone wild-type and histone-mutant tumors. We combine RCAS-H3.3K27M, RCAS-H3.3G34R/V, or RCAS-H3.3WT along with their driver mutations such as RCAS-shp53 and RCAS-PDGFA or RCAS-PDGFB. These tumors are induced in Nestin-positive cells, each in their respective locations found in the human population. Tumors driven by PDGFB have a significantly lower median survival compared to PDGFA-driven tumors and have increased infiltration of lymphocytes and TAMs, specifically inflammatory monocytes. In vitro bone marrow derived monocyte and microglial cultures demonstrate the BMDM population is most responsible for the production of inflammatory chemokines and angiogenic factors in the tumor microenvironment. We performed histological analyses on over 40 human patient samples to determine the role of the stromal population in TAM infiltration. Matched human samples were also utilized for pan-cancer immune profiling with NanoString to further characterize the innate and adaptive immune microenvironments. These analyses indicate DIPG/K27M tumors have a higher immune cell infiltrate compared to G34R/V and histone wildtype tumors. Further, we observe higher infiltration of T-cell populations in pHGGs compared to adult HGGs, suggesting these tumors may be amenable to immunotherapy despite being considered “immune cold.” These studies provide the critical foundation needed for the development of novel therapeutics targeting these tumors.

PDTM-21. MULTI-MODAL PROFILING OF PEDIATRIC HIGH-GRADE GLIOMA SINGLE CELLS USING PATCH-SEQ

Abstract Pediatric and adult high-grade gliomas are characterized by extensive intra-tumoral transcriptional heterogeneity. When measured by single cell RNA sequencing, gliomas reveal themselves as continuums of stemness and differentiation programs with important implications for therapy, but to date this transcriptional information has not been directly linked to physiological behaviors of cells. Recent work from our group establishes the electrical integration of glioma cells into neural circuitry. One subpopulation of glioma cells participates in glutamatergic synaptic communication with neurons, and a distinct subpopulation of cells sense and respond to extracellular potassium flux of neuronal networks by an entirely distinct mechanism. Our data support a model in which both modes of electrical communication are critical to glioma growth, but current associations between the electrophysiological properties of a cell, its transcriptional profile and developmental state are correlational in nature. Patch-seq is needed to clarify the relationship between transcriptional profiles of quiescent/cycling stem-like cells and the observed electrophysiological behaviors. Put more simply, patch-seq will clarify where the synaptically-connected glioma cells exist along a developmental hierarchy. METHODS Here, we adapt a recently described technique called patch-seq to record the electrophysiological profiles of individual pediatric high-grade glioma cells by whole cell patch-clamp and subsequently isolate their mRNA for single cell sequencing by smart-seq2 and analysis using Seurat. In this way, we couple electrophysiological and transcriptomic profiles to unambiguously assign functional identities to cells with transcriptional profiles along a developmental hierarchy. RESULTS We report the successful adaptation of patch-seq for use with patient-derived diffuse intrinsic pontine glioma (DIPG) xenografts in acute brain slice preparations, enabling evaluation of single glioma cells integrated in intact neural circuitry. CONCLUSIONS Data synthesizing the electrophysiological and transcriptomic profiles of single glioma cells in the context of the developmental hierarchy will be presented.